Massive interstitial copy-neutral loss-of-heterozygosity as evidence for cancer being a disease of the DNA-damage response

Yogesh Kumar,Lei Chen,Taobo Hu,Lin Xu,Peng Zhang,Jian-Min Wang,Zhi Xu,Gusheng Tang,Shao Li,Weiqing Wan,Liwei Zhang,Frank W Pun,Peggy Lee,Jianfeng Yang,Hong Xue,Hong Yang Wang ,Suk Ying Tsang ,Wai Sang Poon ,Xiao Hu ,Xueliang Ding ,Siu Kin Ng ,Wai Kin Mat ,Timothy H.y Cheong ,Yang Li ,Jin Fei Chen

doi:10.1186/s12920-015-0104-2

Abstract

BackgroundThe presence of loss-of-heterozygosity (LOH) mutations in cancer cell genomes is commonly encountered. Moreover, the occurrences of LOHs in tumor suppressor genes play important roles in oncogenesis. However, because the causative mechanisms underlying LOH mutations in cancer cells yet remain to be elucidated, enquiry into the nature of these mechanisms based on a comprehensive examination of the characteristics of LOHs in multiple types of cancers has become a necessity.MethodsWe performed next-generation sequencing on inter-Alu sequences of five different types of solid tumors and acute myeloid leukemias, employing the AluScan platform which entailed amplification of such sequences using multiple PCR primers based on the consensus sequences of Alu elements; as well as the whole genome sequences of a lung-to-liver metastatic cancer and a primary liver cancer. Paired-end sequencing reads were aligned to the reference human genome to identify major and minor alleles so that the partition of LOH products between homozygous-major vs. homozygous-minor alleles could be determined at single-base resolution. Strict filtering conditions were employed to avoid false positives. Measurements of LOH occurrences in copy number variation (CNV)-neutral regions were obtained through removal of CNV-associated LOHs.ResultsWe found: (a) average occurrence of copy-neutral LOHs amounting to 6.9 % of heterologous loci in the various cancers; (b) the mainly interstitial nature of the LOHs; and (c) preference for formation of homozygous-major over homozygous-minor, and transitional over transversional, LOHs.ConclusionsThe characteristics of the cancer LOHs, observed in both AluScan and whole genome sequencings, point to the formation of LOHs through repair of double-strand breaks by interhomolog recombination, or gene conversion, as the consequence of a defective DNA-damage response, leading to a unified mechanism for generating the mutations required for oncogenesis as well as the progression of cancer cells.Electronic supplementary materialThe online version of this article (doi:10.1186/s12920-015-0104-2) contains supplementary material, which is available to authorized users.

Highlights

The presence of loss-of-heterozygosity (LOH) mutations in cancer cell genomes is commonly encountered
The results in the tables showed that the heterozygous Mm residues in the genomes of gastric cancers, gliomas, leukemias, liver cancers and primary and secondary lung cancers all displayed exceptionally high mutation rates leading to an all-sample Percentile mutation of Mm residues (RMm) equal to 9.15 % of all Mm residues analyzed, with a great majority of the mutations giving rise to LOH to yield an RLOH of 8.59 %
This question is important to oncogenesis, and to the post-oncogenesis phase with respect to the mutations needed to implement the manifold hallmarks of the neoplastic state, i.e. sustaining proliferative signaling, evading growth suppressors, resisting cell death, enabling replicative immortality, inducing angiogenesis, and activating invasion and metastasis [57], and complex metabolic reprogramming to support rapid growth even under conditions of fluctuating oxygen tension through enhanced glucose uptake, aerobic glycolysis, decreased conversion of pyruvate to acetyl-CoA etc

Summary

Introduction

The presence of loss-of-heterozygosity (LOH) mutations in cancer cell genomes is commonly encountered. Because the causative mechanisms underlying LOH mutations in cancer cells yet remain to be elucidated, enquiry into the nature of these mechanisms based on a comprehensive examination of the characteristics of LOHs in multiple types of cancers has become a necessity. Owing to the importance of LOHs giving rise to loss of major alleles and inactivation of tumor suppressor genes, hitherto investigations of LOHs in cancers have been focused mainly on LOHs that yield homozygous-minor genotypes. In the present study next-generation sequencing was applied to determine at single-base resolution the LOHs in the genomic sequences of various types of cancers, covering sequence regions that have undergone loss of heterozygosity and single nucleotide changes where a heterozygous position has mutated to a homozygous one. Thirty tumor-control pairs of six different types of cancers including glioma (glioblastoma and astroglioma), acute myeloid leukemia, gastric adenocarcinoma, hepatocellular carcinoma, primary lung cancer (pulmonary squamous-cell carcinoma, adenocarcinoma and neuroendocrinal carcinoma), and lung-tobrain metastatic adenocarcinoma were analyzed with the AluScan platform established by our laboratory, based on the capture of ~8–25 Mb/genome of inter-Alu sequences by inter-Alu PCR amplification using multiple consensual Alu sequence-based primers for next-generation sequencing [7]

Methods

Results

Discussion

Conclusion